Nih imagej fiji3/9/2024 Data points outside the whiskers are marked with the plus symbol. The extracted intensity features are shown plotted on the x-axis. ( E, E’) Intensity features box plot comparisons, in which intensity features were extracted using the Extract-ImageProperties_V0.ijm ImageJ/Fiji script. Scale bar was added using ImageJ/Fiji on the right bottom corner of each image. Representative images show differences in mRNA in situ hybridization efficiency and image acquisition settings. ( A’– D’) Confocal maximum intensity projections of c-fos mRNA after in situ hybridization on mouse brain sections, which were acquired by two experimenters (prefrontal cortex and S1 shown here). The representative images show staining variability and different confocal settings for image acquisition (images were acquired at different resolutions). ( A– D) Confocal maximum intensity projections of neurons immunostained for expression of Fos protein in mouse brain sections (prefrontal cortex shown here), which were acquired by four experimenters. Intensity variability in Fos protein immunohistochemistry and mRNA in situ hybridization. Quanty-cFOS facilitates a rapid, accurate and unbiased spatial mapping of neural activity and can also be easily extended to count other types of labelled cells.ĢD automated cell counts Fos protein c-fos mRNA immunohistochemistry in situ hybridization open-source ImageJ/Fiji tool quantitative analysis. Here, we demonstrate the application of the tool in a step-by-step manner, with video tutorials, making it easy for novice users to implement. We validated the tool using data from brain sections in response to somatosensory stimuli in a user-interactive manner. This allows for the overcoming of variations in the data and the deriving of cell counts registered to specific brain areas in a highly time-efficient and reliable manner. ![]() The algorithms compute the intensity cutoff for positive cells on a user-specified number of images and apply this on all the images to process. Here, we describe a novel open-source ImageJ/Fiji tool, called 'Quanty-cFOS', with an easy-to-use, streamlined pipeline for the automated or semi-automated counting of cells positive for the Fos protein and/or c-fos mRNA on images derived from tissue sections. ![]() Quantitatively analyzing the numbers of cells expressing the Fos protein or c-fos mRNA is a major challenge owing to large human bias, subjectivity and variability in baseline and activity-induced expression. Analysis of neural encoding and plasticity processes frequently relies on studying spatial patterns of activity-induced immediate early genes' expression, such as c-fos.
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